Prof Ammu K.

Radhakrishnan
Pathology Division,
School of Medicine, IMU
 Learning Outcomes:
You will be able to:
1. Describe the production of
platelets and leucocytes
2. Name the major growth factors
involved in the production of
platelets and leucocytes

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 Haemopoiesis
1. Erythropoiesis
2. Megakaryopoiesis
3. Leucopoiesis

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 Megakaryopoiesis
Formation of platelets
Development & Maturation
Occurs in bone marrow

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 Platelets: Stages of
Development and Maturation
1. Haemopoietic stem cell
2. Myeloid stem cell
3. Megakaryoblast
4. Pro-megakaryocyte
5. Megakaryocyte
6. Platelets FOUND IN BLOOD

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 Megakaryocyte
Initially a mononuclear cell
Matures by endomitotic
synchronous nuclear replication
DNA replication without cytoplasmic
separation
No daughter cells produced

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 Megakaryocyte
Number of nuclear lobes
increases in multiples of two
There is no cell division
Polyploid cell
Cytoplasmic volumes enlarges
with each cell division
A large cell
50-150 m
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 Bone Marrow
Smear

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 Megakaryocyte
16th - 64th nucleus stage
nuclear replication and cell growth
stops
Microvesicles form
Platelets liberated
Enter marrow sinuses and blood
1 megakaryocyte
4000 platelets
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 Platelets

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 Platelet Production: Control

Thrombopoietin (TPO)
IL-6
IL-3
MK-CSF
Megakaryocyte-colony stimulating
factor

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 Thrombopoietin (TPO)
Megakaryocyte growth and
development factor
MGDF
353 amino acid protein
Encoded by the THPO gene
chromosome 3p27
High homology with erythropoietin
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 Thrombopoietin (TPO)
Produced by
Liver (main source)
Kidneys, testes, brain, bone marrow
stromal cells
Provide primary signals for
megakaryocyte production
Controls platelet production

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 Interleukin-6 (IL-6)
Similar function with TPO
Essential for forming adequate
quantity of platelets
Controls platelet production
Provide primary signals for
megakaryocyte production

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 IL-3 & MK-CSF
Exhibit megakaryocyte stimulating
activity
Promote formation of
megakaryocyte colony

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 Leucopoiesis
Production of leucocytes
White blood cells
Account for <1% of whole blood
Crucial for immune system
Protects against infections
bacteria, viruses, parasites etc.

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 Granulocytes

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 Agranulocytes

10-14 mm

T-, B-lymphocytes
Natural Killer (NK) cell

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 Leucopoiesis
Sites:
Bone marrow (main site)
Thymus
3 major processes:
1. Granulocytopoiesis
2. Monocyte development
3. Lymphopoiesis

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 Leucopoiesis
All leucocytes originate from the
haemopoietic stem cells
Leucocytes can arise from
1. Myeloid stem cells
2. Lymphoid stem cells

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 Haemopoietic Stem Cell

Myeloid Stem Cells Lymphoid Stem Cells

Megakaryocyte GM Progenitor
Progenitor

Proerythroblast Myeloblast Monoblast

Platelets
Erythrocytes Monocyte/
Granulocytes
Prof Ammu K. Radhakrishnan
Macrophage23
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 Haemopoietic Stem Cell

Myeloid Stem Cells Lymphoid Stem Cells

B-lymphocytes Progenitor
Progenitor
T-lymphocytes NK
Progenitor Progenitor
B-Lymphocytes

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T-Lymphocytes
Prof Ammu K. Radhakrishnan
NK cells24
 Terminology
IL
CFU
Interleukin
Colony forming
units SCF
BFU Stem cell factor
Burst forming CSF
units Colony
stimulating
factor

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 Granulocytes: Development
Originate from HSC in the red
bone marrow
Maturation driven by:
Granulocyte-monocyte colony
stimulating factor (GM-CSF)
Granulocyte colony stimulating
factor (G-CSF)

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 Haemopoiesis: Growth Factors and
Development
SCF
SCF
IL-3
Flt-3L
Myeloid IL-6
Stem Cells

Haemopoietic
Stem Cell

SCF
IL-3
Lymphoid
Stem Cells
IL-6
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 Granulocytopoiesis: Growth Factors
and Development
Myeloid
Stem Cells
GM Progenitor
GM-CSF cells

Haemopoietic
Stem Cell Other Progenitor
cells

Lymphoid
Stem Cells

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 Granulocytopoiesis: Growth Factors
and Development

G-CSF
GM-CSF

Myeloid GM Myeloblast
Stem Cells Progenitor
cell

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 Granulocytopoiesis: Growth
Factors and Development

**
Myeloblast Promyelocyte Myelocyte

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 Myelocyte
Precursor/progenitor for all
granulocytes
One for each of the granulocytes
Neutrophil myelocyte
Eosinophil myelocyte
Basophil myelocyte

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 Granulocytopoiesis: Growth
Factors and Development

Myelocyte Metamyelocyte Band Cell

Mature
Granulocyte
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 Granulocytopoiesis
2 steps:
1. Proliferation/Mitotic
2. Differentiation & maturation

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 Granulocytopoiesis:
Proliferation
Rapid mitotic division
Intermediate stages involved:
Myeloblast
Promyelocyte
Myelocyte
Duration
7.5 days

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 Granulocytopoiesis:
Differentiation & Maturation
Post mitosis
Cells undergo maturation and
differentiation to develop characteristic
morphology and feature
Stages involved
Metamyelocyte
Band cell
Duration
6.5 days
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 Neutrophils: Growth Factors and
Development

Myeloblast
Promyelocyte

GM Neutrophil
Myeloid
Progenitor Myelocyte
Stem Cells
cell

Metamyleocyte
Band cell

NEUTROPHIL
G-CSF, GM-CSF
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 Eosinophils: Growth Factors and
Development

Myeloblast
Promyelocyte

GM Eosinophil
Myeloid
Progenitor Myelocyte
Stem Cells
cell

Metamyelocyte
Band cell

EOSINOPHIL
IL-3, IL-5
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 Basophils: Growth Factors and
Development

Myeloblast
Promyelocyte

GM Basophil
Myeloid
Progenitor Myelocyte
Stem Cells
cell

Metamyelocyte
Band cell

BASOPHIL
?
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 Granulocytes: Stages seen
only in bone marrow
1. Haemopoietic stem cells
2. Myeloid stem cells
3. Myeloblasts
4. Promyelocytes
5. Myelocytes
6. Metamyelocytes

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 Granulocytes: Stages seen in
the periphery
1. Blood
Band cells
Mature granulocytes
2. Connective tissues
Mast cells

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 Peripheral Blood

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 Monocytes: Development
Originate from haemopoietic stem
cell in the red bone marrow
Arise from the myeloid stem cell
Maturation driven by:
GM-CSF
Monocyte-colony stimulating factor
(M-CSF)

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 Haemopoiesis: Growth Factors and
Development
Myeloid
Stem Cells
GM Progenitor
GM-CSF cells

Haemopoietic
Stem Cell Other Progenitor
cells

Lymphoid
Stem Cells

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 Monocytes: Growth Factors and
Development

M-CSF
Myeloid GM Monoblast
GM-CSF
Stem Cells Progenitor
cell

Promonocyte
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 Monocytes: Forms only seen
in bone marrow
1. Haemopoietic stem cells
2. Myeloid stem cells
3. Monoblasts
4. Promonocytes

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 Monocytes: Forms seen in
the periphery
1. Blood
Monocytes
2. Tissues
Macrophages
Various types

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 Lymphopoiesis
Types of cell
1. T-lymphocytes
TCR ab T-cells
TCR gd T-cells
2. B-lymphocytes
3. NK cell

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 Lymphopoiesis

Adapted from Ref (3)

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 Lymphocytes: Development

Takes place in:

1. Bone marrow
B-lymphocytes & NK cells
2. Thymus
T-lymphocytes
Maturation driven by cytokines
IL-2, IL-4, IL-6, IL-7
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 Lymphocyte Development

Pluripotent haemopoietic
Lymphoid stem cell
stem cell

Lymphoid
Progenitor

B-lymphocyte
T-lymphocyte
Progenitor
NK Progenitor
(Pro B-cells)
Progenitor (Pro T-cells)
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 T-Lymphocyte: Development

Pro T-cells
CD4- CD8- TCR- CD3-
Double negative (DN)
Maintained by:
IL-6, IL-7

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 T-Lymphocyte: Development
Rearrangement
Migrate to thymus of TCR heavy
chain (b) genes
(Pro T-cells)

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 Immature T-lymphocyte
V, D, J genes rearrange
Form functional VDJ unit
Produce functional b-chain
A productive VDJ rearrangement
required for T-cell survival

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 Immature T-lymphocyte
b-heavy chain
Associates with a surrogate light
chain
Pre-TCR

Pre-TCR+
CD4-CD8- CD3-
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 T-Lymphocyte: Development
Rearrangement
of TCR heavy
Migrate to thymus
chain (b) genes
(Pro T-cells)

IL-2, IL-4

Pre-TCR+
CD4- CD8- CD3- Immature T-lymphocyte
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 Pre-TCR
Required for survival and
proliferation of immature T-cells
Regulates further rearrangement
TCR g genes
1. Stimulate light chain
rearrangements
2. Produce functional a-chain of
TCR
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 TCR
Adapted from
Ref (3)

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 TCR
Functional b- and a- chain
ab-TCR
Signals for expression of
CD4
CD8
CD3

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 T-Lymphocyte: Development

Immature T-lymphocyte
Pre-TCR+ CD4-CD8- CD3

CD3- TCRab+
CD4+ CD8+

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 T-Lymphocyte: Development

TCRab+ CD3+
CD3- TCRab+
CD4+ CD8+
CD4+ CD8+
DOUBLE POSITIVE (DP)
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 Double Positive T-
lymphocytes
Intermediate stage
Present in thymus
Absent in peripheral circulation
Express
ab-TCR, CD3
CD4, CD8
Undergo positive selection
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 Positive Selection
Double positive (DP) become
single positive (SP) cells
1. TCRab+ CD3+ CD4+ CD8-
2. TCRab+ CD3+ CD8+ CD4-
How?
Depends on affinity of TCR to MHC
proteins

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 gd-TCR T-lymphocytes
Function is not clear
Reported to play a critical role:
Immune regulation
Tumour surveillance
Some primary immune responses
Derived and mature from the same
pro-T cells that also generate the
ab-TCR T-cells
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 B-Lymphocyte Subsets

+ Lymph
nodes

16 June 2014 Prof Ammu K Radhakrishnan Adapted from Ref66(3)

 Ig Expression During B-Cell
Development

Nave
B-cell

16 June 2014 Prof Ammu K Radhakrishnan

Adapted from Ref67 (3)
 Checkpoints in Lymphocyte
Maturation

Adapted from Ref (3)

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 NK cell: Development
Originates from lymphoid stem
cell
Process occurs in bone marrow
Development stages not yet fully
elucidated

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 References
1. Abbas & Lichtman; Basic Immunology (4th Edition);
Elsevier Saunders
2. Peakman and Vergani; Basic and Clinical Immunology
(2nd Edition);
Churchill Livingstone
3. Abbas, Lichtman & Pillai; Cellular and Molecular
Immunology (7th Edition); Elsevier Saunders
4. Haematology an illustrated text (2nd edition). Howard
& Hamilton (Churchill Livingstone)
5. Essential Haematology (5th edition)
Hoffbrand and Pettit (Blackwell)

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